No. 3] Proc. Japan Acad., 53, Ser. B (1977) 99
23. Difference o f Chromosome Exchanges Mitomycin C between Normal and
Pericentric mv (9) Cells
By Yukimasa SHIRAISHI
Department of Anatomy, School of Medicine, Kanazawa
Kanazawa 920, Japan
(Communicated by Sajiro MAKING, M. J. A., June 14,
Due to
University,
1977)
Several reports are available for the finding that mitomycin C (MMC) induces non-random chromosome exchanges in human cells (Nowell 1964 ; Cohen and Shaw 1964 ; Br¢ggerand Johansen 1972).
It has been shown that the majority of exchanges occurs preferen- tially between the constitutive heterochromatin of homologous chromosomes, especially in the secondary constriction regions of chromosomes 1, 9 and 16. This may be a result of the association of these chromosome regions during interphase, either through regular somatic pairing (Rao and Nataraj an 1967) or through a particular gathering effect of MMC (Kobayashi 1960). There is little informa- tion on the mitotic feature of the somatic pairing in chromosomes with rearrangements, particulary with pericentric inversions of mam- malian cells. The present study was attempted to test the effect of MMC on cells with pericentric inv (9), one of the polymorphic varia- tions in man (Hansmann 1976; Shiraishi and Makino 1977), and to compare the frequency of MMC-induced exchanges between normal and pericentric inv (9) cells.
Materials and methods. Heparinized blood samples from a normal person and a patient with pericentric inv (9) were cultured for 72 hrs in PHA-containing RPMI 1640 medium supplemented with 15% fetal calf serum at 37°C. The cultures were treated with mito- mycin C (MMC) at concentrations (lx 10-7 g/ml and 5 x 10-7 g/ml ) for the last 24 hrs before the fixation. Colcemide (0.02 ig/ml) was added to the cultures 90 min prior to fixation. Chromosome prepara- tions were processed according to the conventional air-drying method.
The pericentric inv (9) patient, phenotypically normal but having experienced several abortions, was previously described (Shiraishi and Makino 1977).
Results and discussion. The chromosome aberrations observed in this study were of exchange- and break-types. The chromosomes involved in the exchanges were found in most cases associated at the centromere regions. The breaks observed included chromatid- as well
100 Y. SHIRAISHI [Val. 53(B),
Table I. Chromosome aberrations after mitomycin C(MMC)
treatment with
Table II. Number of chromosomes
chromatid exchanges and breaks exposed to 5X10_7 g/ml MMC for
last 24 hrs in cultures
on individual the
Table III. Exchanges between homologous non-homologous chromosomes
and
No. 3] Chromosome Exchanges Affected by Mitomycin C 101 as isochromatid-breaks. The heterochromatic regions of the chromo- somes were particularly affected in the most cases. The number of induced exchanges and breaks observed is higher after treatment with MMC at 5 x 10-7 g/ml than at 1 x 10-7 g/ml (Table I). The num- ber and distribution of exchanges and breaks in the individual chromosomes or groups are shown in Table II. It was noted then
that the majority of exchanges occurred at the regions of the secon- dary constrictions of chromosomes Nos. 1, 9 and 16, or in the so-called C-bands, whereas the distribution of chromosome breaks differed markedly from that of the exchanges. There was an overrepresen- tation of homologous chromosomes in exchanges involving the three chromosomes (Table III). Many homologous chromatid exchanges were also observable in the centrometric heterochromatin segments of chromosomes Nos. 1, 9 and 16 (Fig. la). This agrees with the find- ings of several other reports (Howell 1964; Cohen and Shaw ; BrOgger and Johansen 1972). However, the frequency of the ex-
changes was significantly reduced in normal No. 9 and inverted No. 9 (Table III, Fig. 1b), while in contrast, the frequency of breaks in- creased remarkably in the inverted chromosome No. 9. The break points of breaks in the inverted No. 9 chromosome correspond well to those in exchanges between the normal No. 9 and inverted No. 9 in the same cell from the treated sample. Fig. 1c exhibits breaks in both normal No. 9 chromosome (upper) and inverted chromosome No. 9 (lower) in the same cell, in which break points were identical between the two chromosomes. This seems to suggest that the pair-
Fig. 1. Examples of chromatid exchanges and breaks in chromosome No. 9 induced by mitomycin C, 5x107 -g/ml.
a : Homologous chromatid exchange in normal No. 9 chromosomes. b: Homologous chromatid exchange in normal No. 9 and inverted No. 9 chromosomes. c: Iso- chromatid break in normal No. 9 (upper) and chromatid break in inverted No. 9 (lower). Note the break points identical with those of exchanges in Fig. 1b.
102 Y. SHIRAISHI [Vol. 53(B), ing in the inverted material is rather incomplete because of the absence of homology. The present results agree with the findings reported by De Marco et al. (1975) in Drosophila: and by Heemert (1977) in onion fly. Information is now very limited concerning somatic pairing of chromosomes with rearrangements, such as pen- centric inversions, in mammalian and human cells. Thus, further investigations are requested, in view of the fact that a very similar feature has been provided in both insect and human materials.
Acknowlegements. The author expresses his sincere thanks to Emeritus Prof. Dr. Sajiro Makino, Hokkaido University, for his kind suggestions and reading the manuscript.
This work was supported in part by grants from the Ministry of Education of Japan (No. 854138).
References
Nowell, P. C. (1964) : Exp. Cell Res., 33, 445-449.
Cohen, M. M., and Shaw, M. W. (1964) : J. Cell Biol., 23, 386-395.
Brogger, A., and Johansen, J. (1972) : Chromosoma (Berl.), 38, 95-104.
Rao, R. N., and Nataraj an, A. T. (1967) : Genetics, 57, 821-825.
Kobayashi, J. (1960) : Cytologia, 25, 280-288.
Hansmann, I. (1976) : Human Genet., 31, 247-262.
Shiraishi, Y., and Makino, S. (1977) : Proc. Japan Acad., 53, 79-82.
Marco, A. de, Cozzi, R., Toti, L. (1975) : Genetica (Den Haag), 45, 489-496.
Heemert, C. van (1977) : Chromosoma (Berl.), 5'9, 193-206.
